The objectives of this study are to undertake a clonal analysis of cellular immune response mediated by cytotoxic T cells (CTL) and by natural killer (NK) cell in an autologous human melanoma system and to examine the cellular and molecular basis of cytotoxic unresonsiveness against autologous melanoma cells. We shall examine a hypothesis that cytotoxic unresponsiveness against autologous melanoma cells may result from antigen specific suppression of cytotoxic lymphocytes. To test this hypothesis that in most autologous situations antigen specific cytotoxic cells exist but their effector function is suppressed, cytotoxic lumphocytes will be cloned from responsive or unresponsive lymphocyte populations in the presence of T cell growth factor (TCGF) in soft agar and in liquid microculture under the limiting dilution condition and by employing a Fluorescein activated cell sorter (FACS) equipped with single cell deposition system. TCGF will be produced by PHA induced cultures of human mononuclear cells from spleen and blood. Multiple clones will be expanded in TCGF, analyzed for: cell type (with monoclonal reagents in FACS and with conventional techniques), cytotoxicity, and specificity of cytotoxicity. Cytotoxicity will be assayed in the 3H proline and 51Cr release assays and specificity will be examined in cold target inhibition method. Antigen-specific autoreactive clones will be employed as effector cells in cytotoxicity assays against autologous targets to search for suppressor cells in the unfractionated or fractionated lymphocytes mixed in the assay in appropriate recombination. Such clones will also be used in the search of circulating serum factors as another potential modulator of cellular immunity. Culture supernatants from autologous and allogeneic MLTI will be assayed for TCGF activity (appears to be a prerequisite for T cell response) of the lymphocytes upon such in vitro immunization. Potential inhibitors of TCGF will also be searched in patients' sera. Results of TCGF activities, clonal responsiveness and suppressor activities will be correlated with clinical data to determine their biological significance. Knowledge gained from this study will help in the design of better immunologic monitoring systems and more effective immunotherapy in cancer.